13 Genetics 2-21-11 - Microbial Genetics Chapter Sections...

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Microbial Genetics Chapter Sections 7.1; 7.2; 7.4; 7.5; 9.2; 10.8; 10.9, p 434-435
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Under What Condition is a Gene(s) Under What Condition is a Gene(s) Expressed by a Bacterium? Expressed by a Bacterium? For example, is a gene expressed under: acid conditions or neutral pH conditions? high temperatures or low temperatures? high iron or low iron? How can you determine this?
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Measuring Bacterial Gene Expression Measuring Bacterial Gene Expression One gene at a time Reporter fusions Reverse transcriptase PCR (RT-PCR; qualitative) Real time RT-PCR (quantitative) Multiple genes Microarrays Proteomics
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lacZ lacY lacA β -Galactosidase Lactose permease Transacetylase +1 mRNA Review: Expression of the  Review: Expression of the  lac lac  genes   genes  ( ( lac lac  operon) in   operon) in  E. coli E. coli AUG AUG AUG Stop Stop Stop rbs rbs rbs P DNA Transcription Translatio n protein
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Reporter Fusions (In General) Reporter Fusions (In General) Figure 12.3 Reporter gene Target gene and its promoter Target gene controlled by its own promoter Fuse a reporter gene with the target gene Reporter gene does not contain its own promoter but when fused to target gene, reporter gene controlled by target gene promoter may or may not have its own ribosome binding site (RBS) reporter gene product is easily monitored LacZ (β-galactosidase) cleaves ONPG to produce a colored product GFP (green fluorescent protein) fluoresces
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Operon fusions (or transcriptional fusions) Reporter gene has its own RBS To study transcription
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13 Genetics 2-21-11 - Microbial Genetics Chapter Sections...

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